We used 13-C/1-H double nuclear magnetic resonance to study intracellular polymer formation of hemoglobin in SS, AS and SC erythrocytes. Polymer formation is maximal at complete deoxygenation, decreasing to zero with increasing oxygen saturation. However, polymer can be detected in SS, and to some extent in SC, erythrocytes throughout much of the physiologically relevant oxygen saturation range. The effect of heterogeneity in intracellular hemoglobin concentration in SS and SC erythrocytes, is most apparent at high oxygen saturation: above 85% oxygen saturation, polymer is found predominantly in the subpopulations of denser cells. These experimental results are in good agreement with the theoretical analysis using the intracellular hemoglobin concentration and composition as the primary determinants of polymer formation and provides a basis, in terms of polymer formation, for understanding the pathophysiology of SS and SC disease. We have also shown that for the various other sickle syndromes, polymer formation calculated based on the population average of red cell parameters correlates strongly with the degree of anemia. We are currently using a rapid method for determining the distribution of intracellular hemoglobin concentration to examine in detail the relationship between polymer formation and disease severity for single individuals within each sickle syndrome. In addition, we are examining other causes of cell heterogeneity, including the number of Alpha-globin genes, hemoglobin F and F-cell content. In related studies we are investigating the ontogeny of red cell heterogeneity, especially in the hemoglobinopathies. These studies of intracellular polymer formation can be useful in evaluation or design of therapy. For example, 5-azacytidine treatment increases hemoglobin F production and decreases hemoglobin S and the number of dense cells, reducing the polymerization potential. Studies on non-covalent modifiers of gelation are continuing with efforts to increase the specificity of action based on structural information from X-ray crystallographic analysis.